Effects of pH on plaque forming unit counts and aggregation of MS2 bacteriophage

Area of Science:

• Virology
• Biophysical Chemistry
• Environmental Science

Background:

• Plaque forming unit (PFU) assays are standard for quantifying infectious viral particles.
• Viral particle stability and aggregation are influenced by environmental factors, including pH.
• The isoelectric point (pI) represents the pH at which a particle has no net electrical charge, potentially affecting its stability.

Purpose of the Study:

• To investigate if aggregation processes in aqueous solutions explain the reduction in PFU counts for MS2 phages at pH values near their isoelectric point (pI).

Main Methods:

• Particle size analysis using dynamic light scattering (DLS) and flow particle image analysis.
• Measurement of viral suspensions' aggregate state and size distribution.
• Comparison of PFU counts at different pH values relative to the MS2 phage pI.

Main Results:

• A significant loss in PFU counts was observed for pH values less than or equal to the pI of MS2 phages (pI = 3.9).
• At pH 2.5, a reduction of approximately 3 log(10) PFU was recorded.
• Particle size analysis revealed a shift from approximately 30 nm at neutral pH to several micrometres below the pI, indicating substantial aggregation.

Conclusions:

• MS2 phages undergo significant aggregation at pH values below or equal to their pI, forming aggregates measuring several micrometres.
• This aggregation phenomenon is a primary cause for the observed decline in PFU counts.
• Viral aggregation can introduce considerable bias in PFU assays, necessitating careful control of storage conditions (maintaining pH far from pI) to prevent artefacts.